Typically, skin heals with a scar without regenerating its previous normal structure that includes hair follicles and sweat glands. Scarring causes contractures and morbidity. Impaired healing without skin regeneration leads to chronic wounds. Understanding why skin does not regenerate after wounding would lead to new approaches for treating both acute and chronic wounds, which are major medical problems facing our health system. Hair follicle neogenesis (HFN) after wounding serves as a paradigm for mammalian skin regeneration. We found that following full-thickness excision of mouse skin on the back, hair follicles formed de novo in the healed wound. The skin in the area of hair follicle formation appeared normal without a scar and with the presence of new hair follicles, sebaceous glands and adipose tissue. Based on prior reports, it is possible that similar, albeit much less robust levels of regeneration also occur in humans. The goal of the parent grant is to characterize the cellular and molecular events associated with HFN following wounding. We have studied this remarkable regenerative capacity of the skin using gene expression arrays. Fibroblast growth factor 9 (Fgf9) was upregulated in the epidermis after reepithelialization, but prior to hair germ formation. Using a wounding model for hair follicle neogenesis, we showed that inhibiting Fgf9 resulted in a statistically significant decrease in new hair follicle formation. Over expression of Fgf9 in inducible K14/Fgf9 transgenic mice resulted in marked increases in hair follicle formation during wound healing. Remarkably, we discovered that gamma/delta T cells are the major source of Fgf9 in the epidermis. This finding established a potential connection between skin regeneration and the immune system, and initiated the proposed collaboration between Dr. George Cotsarelis, an expert in epithelial stem cells and PI of the parent grant, and Dr. Avinash Bhandoola, an expert in T cell immunology in the Department of Pathology and Laboratory Medicine at Penn. Epidermal gamma/delta T cells were previously implicated in wound repair in the mouse, but their possible role in hair follicle regeneration has not been studied. Intriguingly, these cells are absent in human epidermis, although they are present in the dermis. Could this explain why human skin regeneration is so severely limited compared to the mouse? We propose to: 1. Determine if hair follicle regeneration occurs in mice lacking gamma/delta T cells. 2. Delete Fgf9 in T cells using Lck-Cre/floxedFgf9 mice to test the effect of loss of Fgf9 on hair follicle neogenesis following wounding, 3. Define differences between different types of gamma/delta T cells in their ability to make Fgf9 and induce hair follicle neogenesis, 4. Determine differences between mouse and human gamma/delta T cells with respect to their wound healing response (ability to produce Fgf9 and other growth factors). These studies lend themselves to interdisciplinary efforts between cutaneous biologists and immunologists that will push research of skin regeneration into new directions. The preliminary results generated here would form a nidus for a larger interactive research project on the role of T cell subsets in skin regeneration and development. Dr. Avinash Bhandoola's expertise in T cell development combined with Dr. Cotsarelis's knowledge of epithelial stem cells and hair biology will provide a strong collaborative effort for the dissection of the role(s) of the immune system in skin regeneration. Ultimately, the findings could impact on medical problems related to wound healing, scar formation, aging and other degenerative conditions.